Experimental Study On Seeding Cells And Vascular Matrix Of Complete Biological Tissue Engineering Blood Vessel

A best substitute of blood vessel has been pursued for a long time and many materials have been tried such as autologous artery or vein,artificial material,biomaterial,but no ideal substitute has been found up till now。Tissue engineering is a new and rapidly expanding field,in which a series of techniques are being developed for culturing and reconstructing a variety of tissues both in vitro and in vivo。With these techniques,it is possible to reconstruct vascular model in vitro whose structure and function are similar to autogenous vessels。In our experiment,endothelial progenitor cells and smooth muscle cells were cultured for the source of seeding cells,Decellularized vascular matrix were obtained from pig aorta by removing its cells,we wish to offer cells and scaffold for further research of TEBV。Objects:1.To provide seeding cells for vascular tissue engineering research;2.To provide decellularized vascular matrix for vascular tissue engineering research。Methods:1.Pig circumference blood was obtained under aseptic condition with heparin。Circumference mononuclear cells were isolated by gradient density centrifugation with lymphocyte separation medium。Endothelial progenitor cells(EPCs) were then obtained and cultured in DMEM。Observed by inverted microscope(IM) and determined by immunohistochemistry(IHC);pig aorta was obtained under aseptic condition with heparin,smooth muscle cells(SMC) were isolated by improved tissue explant method and cultured in DMEM。Observed by inverted microscope (IM) and determined by immunohistochemistry(IHC)。2.Trypsin,hypertonic solutions and chemical detergent were applied for a multi-step process to carry out the preparation of acellular matrix from pig in this study。The specimen was made histology observation investigated its collagen protein level and mechanics character were evaluated。Results:1.EPCs were cultured and sub-cultured successfully,EPCs show "cobblestone" appearance,the CD31,CD34,Flk-1 and the vWF immunity fluorescence dyeing was positive;SMCs were cultured and sub-cultured successfully,and show "hill and vale" appearance under inverted microscope,and a-actin was positive。2.Obtained the decellularized vascular matrix successfully,cells were removed completely,the elastic fibers and collagen fibers were preserved well,the basal membrane is complete。There was no obvious difference in collagen content and mechanics character to the fresh tissue。Conclusion:1.The EPCs and SMCs can be used as the cell source for the vascular tissue engineering;2.The decellularized vascular matrix can be used as the scaffold for the vascular tissue engineering。...